Fibrocytes
Inflammation is the coordinated response to tissue injury or infection. The initiating events are mediated by local release of chemotactic factors, platelet activation, and initiation of the coagulation and complement pathways. These events stimulate the local endothelium, promoting the extravasation of neutrophils and monocytes. The second phase of inflammation is characterized by the influx into the tissue of cells of the adaptive immune system, including lymphocytes. The subsequent resolution phase, when apoptosis of the excess leukocytes and engulfment by tissue macrophages takes place, is also characterized by repair of tissue damage by stromal cells, such as fibroblasts.
Both IL-4 and IL-13 are potent activators of the fibrotic response. IL-4 in known to enhance wound repair and healing. IL-13 has a high degree of homology with IL-4 and in many systems they act in a similar manner. However, key differences have been found in the function of these two proteins in various circumstances. For instance, IL-13 is more dominant in resisting infection by intestinal nematodes and intracellular parasites, such as Leishmania. IL-13 also plays a much more significant role than IL-4 in asthma. In contrast, IL-4 is more dominant than IL-13 in stimulating B cell production of immunoglobulin and in T cell survival and differentiation.
TGFβ, which is also known to play a role in wound healing, had been shown to facilitate fibrocyte differentiation into myofibroblasts, which are further associated with wound healing.
Although IL-4, IL-13, TGFβ and various other factors are known play a role in the fibrotic response, the source of fibroblasts responsible for repair of wound lesions or in other fibrotic responses is controversial. The conventional hypothesis suggests that local quiescent fibroblasts migrate into the affected area, produce extracellular matrix proteins, and promote wound contraction or fibrosis. An alternative hypothesis is that circulating fibroblast precursors (called fibrocytes) present within the blood migrate to the sites of injury or fibrosis, where they differentiate and mediate tissue repair and other fibrotic responses.
Fibrocytes are known to differentiate from a CD14+ peripheral blood monocyte precursor population. Fibrocytes express markers of both hematopoietic cells (CD45, MHC class II, CD34) and stromal cells (collagen types I and III and fibronectin). Mature fibrocytes rapidly enter sites of tissue injury where they secrete inflammatory cytokines. Once there, fibrocytes can function as antigen presenting cells (APCs), thereby inducing antigen-specific immunity. Fibrocytes are also capable of secreting extracellular matrix proteins, cytokines and pro-angiogenic molecules, which may aid in wound repair.
Fibrocytes are also associated with a variety of other processes and disorders. They are associated with the formation of fibrotic lesions after Schistosoma japonicum infection in mice and are also implicated in fibrosis associated with autoimmune diseases. Fibrocytes have also been implicated in pathogenic fibrosis such as that associated with radiation damage, Lyme disease and pulmonary fibrosis. CD34+ fibrocytes have also been associated with stromal remodeling in pancreatitis and stromal fibrosis, whereas lack of such fibrocytes is associated with pancreatic tumors and adenocarcinomas. This correlation may relate to the ability of fibrocytes to function as APCs. Finally, fibrocytes have been shown to promote angiogenesis by acting on endothelial cells.
Serum Amyloid P
Serum amyloid P (SAP), a member of the pentraxin family of proteins that include C-reactive protein (CRP), is secreted by the liver and circulates in the blood as stable pentamers. The exact biological role of SAP is still unclear, although it appears to play a role in both the initiation and resolution phases of the immune response. SAP binds to sugar residues on the surface of bacteria leading to their opsonisation and engulfment. SAP also binds to free DNA and chromatin generated by apoptotic cells at the resolution of an immune response, thus preventing a secondary inflammatory response. Molecules bound by SAP are removed from extracellular areas due to the ability of SAP to bind to all three classical Fcγ receptors (FcγR), with a preference for FcγRI (CD64) and FcγRII (CD32). After receptor binding, SAP and any attached molecule are likely engulfed by the cell.
FcγR are necessary for the binding of IgG to a wide variety of hematopoietic cells. Peripheral blood monocytes express both CD64 and CD32, whereas tissue macrophages express all three classical FcγR. A subpopulation of monocytes also express CD16 (FcγRII).
Clustering of FcγR on monocytes by IgG, either bound to pathogens or as part of an immune complex, initiates a wide variety of biochemical events. The initial events following receptor aggregation include the activation of a series of src kinase proteins. In monocytes, these include lyn, hck and fgr, which phosphorylate tyrosine residues on the ITAM motif of the FcR-γ chain associated with FcγRI and FcγRIII, or the ITAM motif with the cytoplasmic domain of FcγRII. Phosphorylated ITAMs lead to the binding of a second set of src kinases, including syk. Syk has been shown to be vital for phagocytosis of IgG-coated particles. However, the wide distribution of syk in non-hematopoietic cells and the evidence that syk is involved in both integrin and G-protein coupled receptor signaling, indicates that this molecule has many functions.
Both SAP and CRP augment phagocytosis and bind to Fcγ receptors on a variety of cells. CRP binds with a high affinity to FcγRII (CD32), a lower affinity to FcγRI (CD64), but does not bind FcγRIII (CD16). SAP binds to all three classical Fcγreceptors, with a preference for FcγRI and FcγRII, particularly FCγRI. Although there are conflicting observations on the binding of CRP to FcγR, both SAP and CRP have been shown to bind to Fc receptors and initiate intracellular signaling events consistent with FcγR ligation.
In human blood serum, males normally have approximately 32 μg/ml+/−7 μg/ml of SAP, with a range of 12-50 μg/ml being normal. Human females generally have approximately 24 μg/ml+/−8 μg/ml of SAP in blood serum, with a range of 8-55 μg/ml being normal. In human cerebral spinal fluid there is normally approximately 12.8 ng/ml SAP in human males and approximately 8.5 ng/ml in females. Combining male and female data, the normal SAP level in human serum is 26 μg/ml+/−8 μg/ml with a range of 12-55 μg/ml being normal. (The above serum levels are expressed as mean +/−standard deviation.)
SAP has been investigated primarily in relation to its role in amyloidosis. Recently, a drug, R-1-[6-[R-2-carboxy-pyrrolidin-1-yl]-6-oxo-hexanoyl] pyrrolidine-2-carboxylic acid (CPHPC) was developed to deplete SAP and thereby treat amyloidosis. However, this drug appears to have been applied systemically and not to have been used to treat wound healing or to have other localized or systemic effects.
Agar has been previously used as a wound dressing. However, it is not clear whether such previous wound dressings were capable of depleting SAP because they may not have contained appropriate chemical moieties or may have been used inappropriately. In any event, these previous wound dressing do not appear to have incorporated any additional wound healing factors. Further the dressings appear to have been used only for external wounds. Finally, it does not appear that these dressings incorporated purified SAP depleting chemicals or enhanced levels thereof.